Multiple Mitochondrial DNA Deletions Research Articles

P4.61 Prominent respiratory muscle involvement in adPEO with POLG1 mutation

Progressive external ophthalmoplegia (PEO) is characterized by multiple mitochondrial DNA deletions in skeletal muscle. The most common clinical features include adult onset of weakness of the external eye muscles and exercise intolerance. Additional symptoms are variable, and may include cataracts, hearing loss, sensory axonal neuropathy, ataxia, depression, hypogonadism, and parkinsonism. Both autosomal dominant (adPEO) and autosomal recessive inheritance can occur, and autosomal recessive PEO is usually more severe.

POLG1 Variations Presenting as Multiple Sclerosis

To describe 2 unrelated patients with novel variations in the POLG1 gene and features undistinguishable from multiple sclerosis, ie, optic neuritis, brain white matter hyperintense areas, and unmatched cerebrospinal fluid oligoclonal bands. Case report. University hospital. Patients Both patients subsequently developed bilateral ophthalmoplegia, ptosis, myopathy, cardiomyopathy, ataxia, dysphagia, and hearing and cognitive impairment. Detailed clinical and laboratory examinations including brain magnetic resonance imaging, morphological analysis of a muscle biopsy, characterization of mitochondrial DNA integrity, sequencing of the POLG1 gene, and screening of control subjects for POLG1 sequence variants. Ragged red fibers and multiple deletions of mitochondrial DNA were detected in the skeletal muscle. Four compound heterozygous variations, including 3 previously unreported, were identified in POLG1. Clinicians should be aware of the existence of POLG1-related multiple sclerosis-like illness, as it has important implications for management, treatment, and genetic counseling.

Isolated Distal Myopathy of the Upper Limbs Associated With Mitochondrial DNA Depletion and Polymerase γ Mutations

To describe an unusual clinical phenotype in an adult harboring 2 compound heterozygous polymerase γ (POLG) mutations. Case report. University-based outpatient neurology clinic and pathology and genetics laboratory. A 27-year-old man presenting with isolated distal myopathy of the upper extremities in the absence of sensory disturbances. Histochemical analysis of a muscle biopsy specimen showed numerous cytochrome c oxidase-deficient fibers. Molecular analysis revealed marked depletion of muscle mitochondrial DNA in the absence of multiple mitochondrial DNA deletions. Sequence analysis of the POLG gene revealed heterozygous sequence variants in compound c.1156C>T (p.R386C) and c.2794C>T (p.H932Y) segregating with clinical disease in the family. The p.R386C change appears to be a novel mutation. Our case broadens the phenotypic spectrum of disorders associated with POLG mutations and highlights the complex relationship between genotype and phenotype in POLG-related disease.

MELAS syndrome associated with both A3243G-tRNALeu mutation and multiple mitochondrial DNA deletions

The syndrome of mitochondrial encephalopathy, lactic acidosis, and stroke-like episode (MELAS) is characterized clinically by recurrent focal neurological deficits, epilepsy, and short stature. The phenotypic spectrum is extremely diverse, with multisystemic organ involvement leading to isolated diabetes, deafness, renal tubulopathy, hypertrophic cardiomyopathy, and retinitis pigmentosa. In 80% of cases, the syndrome is associated with an AG transmission mutation (A3243G) in the tRNALeu gene of the mitochondrial DNA (mtDNA).We describe a woman with a unique combination of the MELAS A3243G mutation and multiple mtDNA deletions with normal POLG sequence. The patient presented with diabetes mellitus, sensorineural deafness, short stature, and mental disorientation. All her three children died in early adolescence.

Mitochondrial Respiratory Chain Dysfunction in Muscle From Patients With Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is a major cause of neurological disability and its pathogenesis remains elusive despite a multitude of studies. Although defects of the mitochondrial respiratory chain have been described in several ALS patients, their pathogenic significance is unclear. To review systematically the muscle biopsy specimens from patients with typical sporadic ALS to search for possible mitochondrial oxidative impairment. Retrospective histochemical, biochemical, and molecular studies of muscle specimens. Tertiary care university. Subjects Fifty patients with typical sporadic ALS (mean age, 55 years). Main Outcome Measure Number of patients showing a clear muscle mitochondrial dysfunction assessed through histochemical and biochemical muscle analysis. Histochemical data showed cytochrome c oxidase (COX)-negative fibers in 46% patients. Based on COX histochemical activity, patients fell into 4 groups: 27 had normal COX activity; and 8 had mild (2-4 COX-negative fibers of 100 fibers), 8 had moderate (5-10 COX-negative fibers of 100), and 7 had severe (>10 COX-negative fibers of 100) COX deficiency. Spectrophotometric measurement of respiratory chain activities showed that 3 patients with severe histochemical COX deficiency also showed combined enzyme defects. In 1 patient, COX deficiency worsened in a second biopsy taken 9 months after the first. Among the patients with severe COX deficiency, one had a new mutation in the SOD1 gene, another a mutation in the TARDBP gene, and a third patient with biochemically confirmed COX deficiency had multiple mitochondrial DNA deletions detectable by Southern blot analysis. Our data confirm that the histochemical finding of COX-negative fibers is common in skeletal muscle from patients with sporadic ALS. We did not find a correlation between severity of the oxidative defect and age of the patients or duration of the disease. However, the only patient who underwent a second muscle biopsy did show a correlation between severity of symptoms and worsening of the respiratory chain defect. In 7 patients, the oxidative defect was severe enough to support the hypothesis that mitochondrial dysfunction must play a role in the pathogenesis of the disease.

OPA1 mutations cause cytochrome c oxidase deficiency due to loss of wild-type mtDNA molecules

Pathogenic OPA1 mutations cause autosomal dominant optic atrophy (DOA), a condition characterized by the preferential loss of retinal ganglion cells and progressive optic nerve degeneration. Approximately 20% of affected patients will also develop more severe neuromuscular complications, an important disease subgroup known as DOA+. Cytochrome c oxidase (COX)-negative fibres and multiple mitochondrial DNA (mtDNA) deletions have been identified in skeletal muscle biopsies from patients manifesting both the pure and syndromal variants, raising the possibility that the accumulation of somatic mtDNA defects contribute to the disease process. In this study, we investigated the mtDNA changes induced by OPA1 mutations in skeletal muscle biopsies from 15 patients with both pure DOA and DOA+ phenotypes. We observed a 2- to 4-fold increase in mtDNA copy number at the single-fibre level, and patients with DOA+ features had significantly greater mtDNA proliferation in their COX-negative skeletal muscle fibres compared with patients with isolated optic neuropathy. Low levels of wild-type mtDNA molecules were present in COX-deficient muscle fibres from both pure DOA and DOA+ patients, implicating haplo-insufficiency as the mechanism responsible for the biochemical defect. Our findings are consistent with the ‘maintenance of wild-type’ hypothesis, the secondary mtDNA deletions induced by OPA1 mutations triggering a compensatory mitochondrial proliferative response in order to maintain an optimal level of wild-type mtDNA genomes. However, when deletion levels reach a critical level, further mitochondrial proliferation leads to replication of the mutant species at the expense of wild-type mtDNA, resulting in the loss of respiratory chain COX activity.

The clinical, histochemical, and molecular spectrum of PEO1 (Twinkle)-linked adPEO

Mutations in the Twinkle (PEO1) gene are a recognized cause of autosomal dominant progressive external ophthalmoplegia (adPEO), resulting in the accumulation of multiple mitochondrial DNA (mtDNA) deletions and cytochrome c oxidase (COX)-deficient fibers in skeletal muscle secondary to a disorder of mtDNA maintenance. Patients typically present with isolated extraocular muscle involvement, with little apparent evidence of the clinical heterogeneity documented in other mtDNA maintenance disorders, in particular POLG-related disease. We reviewed the clinical, histochemical, and molecular genetics analysis of 33 unreported patients from 26 families together with all previous cases described in the literature to define the clinical phenotype associated with PEO1 mutations. Ptosis and ophthalmoparesis were almost universal clinical features among this cohort, with 52% (17/33) reporting fatigue and 33% (11/33) having mild proximal myopathy. Features consistent with CNS involvement were rarely described; however, in 24% (8/33) of the patients, cardiac abnormalities were reported. Mitochondrial histochemical changes observed in muscle showed remarkable variability, as did the secondary mtDNA deletions, which in some patients were only detected by PCR-based assays and not Southern blotting. Moreover, we report 7 novel PEO1 variants. Our data suggest a shared clinical phenotype with variable mild multiorgan involvement, and that the contribution of PEO1 mutations as a cause of adPEO may well be underestimated. Direct sequencing of the PEO1 gene should be considered in adPEO patients prior to muscle biopsy.

Polyneuropathy as a multisystemic plus symptom in chronic progressive external ophthalmoplegia: Phenotypic differences in between singular deletions and multiple deletions of mitochondrial DNA

Polyneuropathy as a multisystemic plus symptom in chronic progressive external ophthalmoplegia: Phenotypic differences in between singular deletions and multiple deletions of mitochondrial DNA

Multi-system neurological disease is common in patients with OPA1 mutations

Additional neurological features have recently been described in seven families transmitting pathogenic mutations in OPA1, the most common cause of autosomal dominant optic atrophy. However, the frequency of these syndromal ‘dominant optic atrophy plus’ variants and the extent of neurological involvement have not been established. In this large multi-centre study of 104 patients from 45 independent families, including 60 new cases, we show that extra-ocular neurological complications are common in OPA1 disease, and affect up to 20% of all mutational carriers. Bilateral sensorineural deafness beginning in late childhood and early adulthood was a prominent manifestation, followed by a combination of ataxia, myopathy, peripheral neuropathy and progressive external ophthalmoplegia from the third decade of life onwards. We also identified novel clinical presentations with spastic paraparesis mimicking hereditary spastic paraplegia, and a multiple sclerosis-like illness. In contrast to initial reports, multi-system neurological disease was associated with all mutational subtypes, although there was an increased risk with missense mutations [odds ratio = 3.06, 95% confidence interval = 1.44–6.49; P = 0.0027], and mutations located within the guanosine triphosphate-ase region (odds ratio = 2.29, 95% confidence interval = 1.08–4.82; P = 0.0271). Histochemical and molecular characterization of skeletal muscle biopsies revealed the presence of cytochrome c oxidase-deficient fibres and multiple mitochondrial DNA deletions in the majority of patients harbouring OPA1 mutations, even in those with isolated optic nerve involvement. However, the cytochrome c oxidase-deficient load was over four times higher in the dominant optic atrophy + group compared to the pure optic neuropathy group, implicating a causal role for these secondary mitochondrial DNA defects in disease pathophysiology. Individuals with dominant optic atrophy plus phenotypes also had significantly worse visual outcomes, and careful surveillance is therefore mandatory to optimize the detection and management of neurological disability in a group of patients who already have significant visual impairment.

A new locus on 3p23–p25 for an autosomal-dominant limb-girdle muscular dystrophy, LGMD1H

Limb-girdle muscular dystrophies (LGMDs) are a genetically heterogeneous group of neuromuscular disorders with a selective or predominant involvement of shoulder and pelvic girdles. We clinically examined 19 members in a four-generation Italian family with autosomal-dominant LGMD. A total of 11 subjects were affected. Clinical findings showed variable expressivity in terms of age at onset and disease severity. Five subjects presented with a slowly progressive proximal muscle weakness, in both upper and lower limbs, with onset during the fourth-fifth decade of life, which fulfilled the consensus diagnostic criteria for LGMD. Earlier onset of the disease was observed in a group of patients presenting with muscle weakness and/or calf hypertrophy, and/or occasionally high CK and lactate serum levels. Two muscle biopsies showed morphological findings compatible with MD associated with subsarcolemmal accumulation of mitochondria and the presence of multiple mitochondrial DNA deletions. A genome-wide scan performed using microsatellite markers mapped the disease on chromosome 3p23-p25.1 locus in a 25-cM region between markers D3S1263 and D3S3685. The highest two-point LOD score was 3.26 (theta=0) at marker D3S1286 and D3S3613, whereas non-parametric analysis reached a P-value=0.0004. Four candidate genes within the refined region were analysed but did not reveal any mutations. Our findings further expand the clinical and genetic heterogeneity of LGMDs.

Novel Twinkle gene mutation in autosomal dominant progressive external ophthalmoplegia and multisystem failure

A Saudi Arabian family presented with adult onset autosomal dominant progressive external ophthalmoplegia (adPEO) complicated by late onset reversible failure of the CNS, respiratory, hepatic, and endocrine systems. Clinical findings were suggestive of mitochondrial dysfunction and multiple mitochondrial DNA deletions were demonstrated on long range and real time polymerase chain reaction assays but not on Southern blotting. The disorder is caused by a novel heterozygous PEO1 mutation predicting a Leu360Gly substitution in the twinkle protein. The peculiar clinical presentation expands the variable phenotype observed in adPEO and Twinkle gene mutations.

Clinical phenotype of autosomal dominant progressive external ophthalmoplegia in a family with a novel mutation in the C10orf2 gene

Autosomal dominant progressive external ophthalmoplegia (adPEO) is a mitochondrial disorder caused by mutations in nuclear genes. Here we report the clinical and genetic features of adPEO in a Chinese family. All patients had gradual onset of ptosis, with or without ophthalmoplegia, around age 30. Thirteen patients had limb weakness around age 40. Eight patients developed dysphagia around age 50. Four patients died of cardiac abnormalities around age 60. Muscle biopsy of the proband indicated mitochondrial myopathy characterized by ragged-red fibers, cytochrome c oxidase-negative fibers, and multiple deletions of mitochondrial DNA. A heterozygous missense mutation of c.1342A>G in the C10orf2 gene resulting in the p.448N>D mutation in the protein was found in the proband and four other affected family members. In summary, we identified an adPEO family with a novel C10orf2 gene mutation that manifested an age-dependent phenotype. It suggests that greater attention must be paid to cardiac abnormalities in the late stages of this disease.

A Heterozygous Truncating Mutation in RRM2B Causes Autosomal-Dominant Progressive External Ophthalmoplegia with Multiple mtDNA Deletions

Autosomal-dominant progressive external ophthalmoplegia (adPEO) is a mitochondrial disorder that is characterized by accumulation of multiple mitochondrial DNA (mtDNA) deletions in postmitotic tissues. The disorder is heterogeneous, with five known nuclear disease genes that encode the proteins ANT1, Twinkle, POLG, POLG2, and OPA1. Defects in these proteins affect mtDNA maintenance, probably leading to stalled replication forks, consequent mtDNA deletion formation, and progressive respiratory chain deficiency. Here we present a large adPEO family with multiple mtDNA deletions, whose disease was not explained by mutations in any of the known adPEO loci. We mapped the disease locus in this family to chromosome 8q22.1-q23.3. The critical linkage region contained the RRM2B gene, which encodes the small subunit of the ribonucleotide reductase p53R2, which has previously been shown to be essential for the maintenance of mtDNA copy number. Mutation screening of RRM2B revealed a heterozygous nonsense mutation in exon 9 (c.979C-->T [p.R327X]) in all affected individuals that was absent in 380 control chromosomes. The same mutation was found to segregate in another adPEO family. The mutant mRNA escaped nonsense-mediated decay and resulted in a protein with truncation of 25 highly conserved C-terminal amino acids essential for the interaction with the ribonucleotide reductase subunit R1. We conclude that dominant-negative or gain-of-function mutations in RRM2B are a cause of multiple mtDNA deletions and adPEO.

Secondary mtDNA Defects Do Not Cause Optic Nerve Dysfunction in a Mouse Model of Dominant Optic Atrophy

The majority of patients with autosomal dominant optic atrophy (DOA) harbor pathogenic OPA1 mutations and certain missense mutations, mostly within the GTPase domain, have recently been shown to cause multiple mitochondrial DNA (mtDNA) deletions in skeletal muscle. This raises the possibility that the optic neuropathy could be the result of secondary mtDNA defects accumulating within retinal ganglion cells (RGCs). To explore this hypothesis, the authors looked for evidence of mitochondrial dysfunction in a mouse model of DOA and documented the visual and neurologic progression in aging mutant mice. Visual function was assessed with a rotating optokinetic (OKN) drum at ages 13 and 18 months and neurologic phenotyping was performed using the primary SHIRPA screen at age 13 months, comparing mutant Opa1(+/)(-) mice with wild-type C57Bl/6 mice. The presence of cytochrome c oxidase (COX) deficiency and multiple mtDNA deletions was investigated in gastrocnemius muscle and eye specimens harvested from 2- and 11-month-old Opa1(+/+) and Opa1(+/)(-) mice. At age 13 months, Opa1(+/)(-) mice had a statistically significant reduction in OKN responses compared to C57Bl/6 controls with both 2 degrees and 8 degrees gratings (P < 0.001). At age 18 months, the difference between the two groups was significant for the 8 degrees grating (P = 0.003) but not for the 2 degrees grating (P = 0.082). Opa1(+/)(-) mice did not exhibit any significant neuromuscular deficits and no COX deficient areas or secondary mtDNA deletions were identified in skeletal muscle or the RGC layer. There was also no evidence of significant mtDNA depletion or proliferation in skeletal muscle from Opa1(+/)(-) mice. COX deficiency and mtDNA abnormalities do not contribute to optic nerve dysfunction in pure DOA.

Molecular analysis in a family presenting with a mild form of late-onset autosomal dominant chronic progressive external ophthalmoplegia

Nuclear genes affecting mitochondrial genome stability were screened in an Italian family presenting with autosomal dominant progressive external ophthalmoplegia (adPEO) associated with multiple mitochondrial DNA (mtDNA) deletions. We report on a heterozygous c.907C>T (p.R303W) mutation found in the N-terminal domain of the human mitochondrial DNA helicase, Twinkle protein, in six members of a family, in which two individuals manifested late-onset PEO and morphological and molecular signs of mitochondrial dysfunction along with two carriers who are presently free of disease manifestation. We also investigated if the p.R303W mutation in PEO1 gene affected the relative copy number of mitochondrial DNA genomes.

Finding twinkle in the eyes of a 71‐year‐old lady: A case report and review of the genotypic and phenotypic spectrum of TWINKLE‐related dominant disease

Progressive external ophthalmoplegia (PEO) can be caused by a disorder characterized by multiple mitochondrial DNA (mtDNA) deletions due to mutations in the TWINKLE gene, encoding a mtDNA helicase. We describe a 71-year-old woman who had developed PEO at age 55 years. She had cataracts, diabetes, paresthesias, cognitive defects, memory problems, hearing loss, and sensory ataxia. She had muscle weakness with ragged red fibers on biopsy. MRI showed static white matter changes. A c.908G>A substitution (p.R303Q) in the TWINKLE gene was identified. Multiple mtDNA deletions were detected in muscle but not blood by a PCR-based method, but not by Southern blot analysis. MtDNA copy number was maintained in blood and muscle. A systematic literature search was used to identify the genotypic and phenotypic spectrum of dominant TWINKLE-related disease. Patients were adults with PEO and symptoms including myopathy, neuropathy, dysarthria or dysphagia, sensory ataxia, and parkinsonism. Diabetes, cataract, memory loss, hearing loss, and cardiac problems were infrequent. All reported mutations clustered between amino acids 303 and 508 with no mutations at the N-terminal half of the gene. The TWINKLE gene should be analyzed in adults with PEO even in the absence of mtDNA deletions in muscle on Southern blot analysis, and of a family history for PEO. The pathogenic mutations identified 5' beyond the linker region suggest a functional role for this part of the protein despite the absence of a primase function in humans. In our patient, the pathogenesis involved multiple mtDNA deletions without reduction in mtDNA copy number.

Recessive twinkle mutations cause severe epileptic encephalopathy

The C10orf2 gene encodes the mitochondrial DNA helicase Twinkle, which is one of the proteins important for mitochondrial DNA maintenance. Dominant mutations cause multiple mitochondrial DNA deletions and progressive external ophthalmoplegia, but recent findings associate recessive mutations with mitochondrial DNA depletion and encephalopathy or hepatoencephalopathy. The latter clinical phenotypes resemble those associated with recessive POLG1 mutations. We have previously described patients with infantile onset spinocerebellar ataxia (MIM271245) caused either by homozygous (Y508C) or compound heterozygous (Y508C and A318T) Twinkle mutations. Our earlier reports focused on the spinocerebellar degeneration, but the 20-year follow-up of 23 patients has shown that refractory status epilepticus, migraine-like headaches and severe psychiatric symptoms are also pathognomonic for the disease. All adolescent patients have experienced phases of severe migraine, and seven patients had antipsychotic medication. Epilepsia partialis continua occurred in 15 patients leading to generalized epileptic statuses in 13 of them. Eight of these patients have died. Valproate treatment was initiated on two patients, but had to be discontinued because of a severe elevation of liver enzymes. The patients recovered, and we have not used valproate in infantile onset spinocerebellar ataxia since. The first status epilepticus manifested between 15 and 34 years of age in the homozygotes, and at 2 and 4 years in the compound heterozygotes. The epileptic statuses lasted from several days to weeks. Focal, stroke-like lesions were seen in magnetic resonance imaging, but in infantile onset spinocerebellar ataxia these lesions showed no predilection. They varied from resolving small cortical to large hemispheric oedematous lesions, which reached from cerebral cortex to basal ganglia and thalamus and caused permanent necrotic damage and brain atrophy. Brain atrophy with focal laminar cortical necrosis and hippocampal damage was confirmed on neuropathological examination. The objective of our study was to describe the development and progression of encephalopathy in infantile onset spinocerebellar ataxia syndrome, and compare the pathognomonic features with those in other mitochondrial encephalopathies.

Mitochondrial disorder with OPA1 mutation lacking optic atrophy

OPA1 is highly expressed in retina and optic nerve. OPA1 mutations were first identified in patients with non-syndromic autosomal dominant optic atrophy. Recently, OPA1 mutations were detected in a multisystemic disorder which has optic atrophy as the core clinical feature and multiple mitochondrial DNA (mtDNA) deletions in muscle. We report a patient with a multisystemic disorder and multiple muscle mtDNA deletions, carrying an in-frame deletion in OPA1 in the absence of optic atrophy. This patient provides evidence that optic atrophy is not the main clinical manifestation of OPA1-related disorders. OPA1 analysis should be considered in mitochondrial disorders despite the lack of optic atrophy.

Phenotypic Variations in 3 Children With POLG1 Mutations

Autosomal inherited mitochondrial diseases have been of increasing interest among clinicians and mitochondrial research groups because these diseases are caused through a secondary effect on the mitochondrial DNA. It was thought that the genetic stability of mitochondrial DNA relies on the accuracy of DNA polymerase gamma. Mutations of DNA polymerase gamma 1 gene (MIM# 174763) have been shown to be a cause of mitochondrial disorders associated with Mendelian disorders characterized by multiple mitochondrial DNA deletions or depletions. To date, several clinical phenotypes associated with polymerase gamma mutation have been reported presenting in both adults and children. We present 3 children in whom were found to have reported pathogenic DNA polymerase gamma 1 mutations: heterozygous p.G517V in 2 half siblings and heterozygous p.T251I and p.P587L in the other. The aim of this communication is to report 3 pediatric cases associated with DNA polymerase gamma 1 mutations to augment the expanding clinical phenotype that has been previously reported.

Alpha‐synuclein pathology and Parkinsonism associated with POLG1 mutations and multiple mitochondrial DNA deletions

Alpha‐synuclein pathology and Parkinsonism associated with <i>POLG1</i> mutations and multiple mitochondrial DNA deletions